ELISA and Immunoassays

Introduction

• Immunoassay — An immunoassay is an analytical method that measures the concentration of an analyte (antigen or antibody) by exploiting the highly specific, high-affinity antigen–antibody (Ag–Ab) binding reaction, with the bound complex reported through a measurable signal (enzyme, radioisotope, fluorophore or electrochemiluminescent label).
• ELISA — ELISA (Enzyme-Linked ImmunoSorbent Assay) is the immunoassay subclass in which the reporter is an enzyme covalently conjugated to one of the immunoreactants; the enzyme converts a colourless chromogenic substrate into a coloured (or luminescent/fluorescent) product whose intensity is proportional to the bound analyte.
• Ligand-binding assay — ELISA is the dominant member of the ligand-binding assay (LBA) family in bioanalysis — the platform that measures large-molecule (biologic) drugs, hormones, cytokines and anti-drug antibodies in biological matrices, distinct from chromatographic assays (HPLC, LC-MS/MS).
• Why it is the workhorse — High specificity (antibody-driven), high sensitivity (enzymatic signal amplification), microvolume (µL-scale) samples, 96-/384-well microtitre-plate throughput, and a non-radioactive readout make ELISA the workhorse of pharmacology bench-work and clinical drug development.